Vagnarelli P, Giulotto E, Fattorini P, Mucciolo E, Bensi M, Tessera L, De Carli L
Dipartimento di Genetica e Microbiologia, Università di Pavia, Italy.
EXS. 1993;67:71-7. doi: 10.1007/978-3-0348-8583-6_7.
A mutation assay in cultured mammalian cells was developed based on direct analysis of minisatellite DNA. Chinese hamster cells (V79) were mutagenized with nitrosoguanidine and independent colonies were isolated and expanded. DNA fingerprints were then obtained after digestion with HinfI or HaeIII and hybridization with 33.15 and 33.6 probes (Jeffreys et al., 1985). 12 colonies from untreated cells were also analyzed. Digestion with HaeIII and hybridization with 33.15 probe detected the highest frequency of induced variants. The results suggest that minisatellite sequences are hypermutable sites that can be used to monitor the mutagenic effect of chemical agents. We have also analyzed the DNA fingerprints of 17 independent Chinese hamster (CHO) cell lines carrying amplification of the CAD gene. The DNA fingerprint analysis showed a variation in minisatellite regions in 3 lines while no variation was observed in independent colonies from the CHO parental cell line. The results suggest that these sequences may be hot spots for recombination during gene amplification.
基于对小卫星DNA的直接分析，开发了一种在培养的哺乳动物细胞中进行的突变检测方法。用亚硝基胍诱变中国仓鼠细胞（V79），分离并扩增独立的菌落。然后用HinfI或HaeIII消化并用33.15和33.6探针杂交（Jeffreys等人，1985年）获得DNA指纹。还分析了12个未处理细胞的菌落。用HaeIII消化并用33.15探针杂交检测到诱导变体的最高频率。结果表明，小卫星序列是高变位点，可用于监测化学试剂的诱变作用。我们还分析了17个携带CAD基因扩增的独立中国仓鼠（CHO）细胞系的DNA指纹。DNA指纹分析显示3个细胞系的小卫星区域存在变异，而CHO亲本细胞系的独立菌落未观察到变异。结果表明，这些序列可能是基因扩增过程中重组的热点。
